Stimulation of THP-1 Macrophages with LPS Increased the Production of Osteopontin-Encapsulating Exosome

• Published in: International journal of molecular sciences 2020-11, Vol.21 (22), p.8490
• Main Authors: Bai, Gaowa, Matsuba, Takashi, Niki, Toshiro, Hattori, Toshio
• Format: Article
• Language: English
• Subjects: Acetic acid; Antibodies; Apoptosis; Binding sites; Biomedical materials; Bone remodeling; Cancer; CD9 antigen; Cell adhesion & migration; Cell Differentiation - drug effects; Cell Line; Encapsulation; Exo-Prep; exosome; Exosomes; Exosomes - drug effects; Humans; Lipopolysaccharides; Lipopolysaccharides - pharmacology; LPS; Macrophages; Macrophages - drug effects; Macrophages - metabolism; Neutrophils; Osteopontin; Osteopontin - metabolism; Phorbol 12-myristate 13-acetate; Phosphorylcholine - analogs & derivatives; Phosphorylcholine - chemistry; Polymethacrylic Acids - chemistry; Stimulation; THP-1; THP-1 Cells - drug effects; THP-1 Cells - metabolism; Tuberculosis; Western blotting
• ISSN: 1422-0067; 1661-6596; 1422-0067
• DOI: 10.3390/ijms21228490

Osteopontin (OPN) mediates bone remodeling and tissue debridement. The OPN protein is cleaved, but it is unclear how full-length (FL)-OPN or its cleaved form perform their biological activities in target cells. We, therefore, performed the molecular characterization of OPN in exosomes (Exo). The Exo were isolated from lipopolysaccharide (LPS)-stimulated phorbol 12-myristate 13-acetate (PMA)-differentiated THP-1 macrophages. The Exo were also isolated from PMA-differentiated THP-1 macrophages. The Exo were identified using the qNano multiple analyzer (diameter 59-315 nm) and western blotting with a CD9 antibody. LPS-stimulated cells produced more particles than non-stimulated cells. The presence of the FL or the cleaved form of OPN was confirmed using western blot analysis. A mixture of FL and cleaved OPN was also measured using an ELISA system (Ud-OPN) and their presence in the Exo was confirmed. Ud/FL ratios became low after LPS stimulation, indicating the enhanced encapsulation of FL-OPN in the Exo by LPS. These findings suggest that LPS stimulation of human macrophages facilitates the synthesis of FL-OPN, which is cleaved in cells or the Exo after release. These findings indicate that Exo is a suitable vehicle to transfer OPN to the target cells.